The Jiangnan Orogen is China’s third largest gold province, and has undergone complex orogenic processes and tectonic overprinting, forming multistage magmatism, deformation, metamorphism, and gold mineralization. Our field studies indicate that the Tuanshanbei gold ores (central Jiangnan Orogen) have two generations of auriferous quartz-ankerite-pyrite-arsenopyrite veins (Q2 and Q3), with the latter containing abundant ankerite and base metal sulfides. Both Q2 and Q3 veins are younger than the emplacement of the Tuanshanbei granodiorite vein-ore host, and cut by post-ore diabase dikes. Q2 veins were likely associated with the Early Devonian near N-S-directed shortening, along sub-horizontal EW-/WNW-striking transpressive faults, whereas Q3 veins (hosting ∼ 70% of the total Au resource), were primarily hosted in the Early Triassic moderately-/steeply-dipping NW-striking tensional/transtensional faults and moderately-dipping NE-/NNE-striking transpressive faults, associated with NW-SE-directed shortening. Discrete structural styles and mineral assemblages from Q2 and Q3 suggest superimposing gold mineralization at Tuanshanbei. To determine the source(s) of the ore-forming aqueous-carbonic fluids, we conducted mineralogical and in-situ LA-(MC)-ICP-MS Nd-isotope analyses on two generations of hydrothermal monazite, and H-O isotope analyses on the quartz from Q2 and Q3 veins.The results suggest that the Tuanshanbei ore fluids have δD = −66.7 to −57.6‰ (Q2) and −66.8 to −66.2‰ (Q3), and calculated δ18OH2O = 4.5–9.7‰ (Q2) and 6.1–7.8‰ (Q3). The H-O isotopic data suggest a metamorphic and/or magmatic water source for the ore fluids. Hydrothermal monazite (coexists with native gold and auriferous sulfides) from Q2 and Q3 veins displays subtle heterogeneous BSE response along cracks and grain margin, suggesting alteration there. The 147Sm/144Nd and 143Nd/144Nd ratios of altered parts of Q2 and Q3 monazite grain are significantly perturbed via dissolution–recrystallization by the hydrothermal fluids, whereas their initial Nd isotope compositions seem to be less affected. The unaltered Q2 and Q3 monazite samples have similar initial εNd values (Q2: −8.07 to −7.36, Q3: −10.87 to −9.99) to the Neoproterozoic Cangxiyan Group greenschist–amphibolite-facies metamorphic rocks. Geological and structural evidence suggest both Q2 and Q3 ore-forming events were related to metamorphism, typical of orogenic gold deposits. We interpreted that various parts of the basement were metamorphosed (to near the greenschist-amphibolite-facies boundary) at different times, during which the gold-bearing metamorphic fluids produced migrated into the same structural conduits and deposited the ores there in two ore-forming episodes.